Complete mitochondrial genome and phylogenetic analysis of the caddisfly Neucentropus mandjuricus (Trichoptera: Polycentropodidae)

Abstract We have sequenced and annotated the complete mitogenome of Neucentropus mandjuricus Martynov, 1907, a monotypic genus of Polycentropodidae with controversial taxonomic status. The complete mitogenome of N. mandjuricus is 15,020 bp in length, including 13 protein-coding genes, two ribosomal genes, 22 transfer RNA genes, and a non-coding control region. The maximum-likelihood and Bayesian’s inference trees based on 13 protein-coding genes of 23 trichopteran species show that N. mandjuricus clusters together with the other polycentropodid Plectrocnemia sp. with high support value. This study provides a basis for further study on mitogenome and phylogenetics of the Polycentropodidae.


Introduction
The Polycentropodidae is a large family among Trichoptera and distributed worldwide, consisting of 30 genera in three subfamilies (Morse 2022). Neucentropus Martynov, 1907 is a monotypic genus of Polycentropodidae, including only one species: Neucentropus mandjuricus Martynov, 1907. However, the taxonomy status of Neucentropus has been controversial as the intraspecific variations widely existed in the wing and external genitalia (Li et al. 1998). The phylogenetic analysis based on three protein-coding genes indicated that Neucentropus combines with Neureclipsis and locates in the most basal branch of Polycentropodidae (Johanson et al. 2012). In this study, we have sequenced and annotated the complete mitogenome of Neucentropus mandjuricus, aiming to provide more molecular data for phylogenetic analysis of Polycentropodidae.

Results and discussion
The complete mitogenome of N. mandjuricus is 15,020 bp in length, with double circular strands consisting of 13 proteincoding genes (PCGs), two ribosomal genes (rRNA), 22 transfer RNA (tRNA) genes, and a non-coding control region ( Figure  2). The mitogenome exhibits AT-bias, with the AT content of 81% (A ¼ 37%, T ¼ 44%, C ¼ 6.3%, G ¼ 12.6%). The total length of 13 PCGs is 10,922 bp, accounting for 72.72% of the whole genome. All PCGs are initiated with ATN codons, as four PCGs (nad3, nad4L, nad5, and nad6) with ATT, four (atp6, cytb, nad1, and nad4) with ATG, four (nad2, cox1, cox3, and atp8) with ATA, and cox2 with ATC. Nad2, cox1, and cox2 use the truncated stop codon (T), while the other PCGs all end with TAA. The 22 tRNA genes vary from 58 bp (trnS) to 72 bp (trnK and trnL). Two rRNA genes (rrn12 and rrn16), located at trnF/control region and trnV/nad1, are 622 bp and 1368 bp in length, respectively. Comparative mitogenome analyses between N. mandjuricus and Plectrocnemia sp. (another Polycentropodinae with complete mitogenome available at GenBank under accession number MW413804) reveals significant collinearity for 13 PCGs. Gene rearrangement is also found for some tRNA genes: trnG is located between trnQ and trnL in N. mandjuricus, whereas in Plectrocnemia sp. it is between trnH and control region.
For phylogenetic analysis, the coding sequences of 13 PCGs from 23 Trichoptera, along with a Lepidoptera species Thitarodes damxungensis (Yang, 1995) as an outgroup, were individually aligned based on the codons using PAL2NAL v14 (Suyama et al. 2006), and concatenated to form a single dataset. The maximum-likelihood (ML) and Bayesian's inference (BI) trees were then generated by RAxML-NG v1.1.0 (Kozlov et al. 2019) andMrBayes v3.2.7 (Ronquist et al. 2012), respectively ( Figure 3). The result shows that N. mandjuricus forms the sister group to Plectrocnemia sp. with high support value (BS ¼ 100, PP ¼ 1). Polycentropodidae and Stenopsychidae have close relationship, forming the sister group to other Annulipalpia. Our study provides more molecular data for further research on evolutionary relationships of Polycentropodidae.

Author contributions
Conceived and designed the study: Chao Yue. Performed the experiments: Zhen Li. Analyzed the data: Na Ma and Ting Lei. Draft the manuscript: Na Ma and Chao Yue. Revised the manuscript: Ting Lei. Final approval of the version to be published: Ting Lei and Chao Yue. All authors agreed to be accountable for all aspects of the work.

Ethics statement
The collected insect samples are not protected by regulations or law and not required an ethical review by the Statement Animal Experiment Committee.